A Kalirin missense mutation enhances dendritic RhoA signaling and leads to regression of cortical dendritic arbors across development.

Normally, dendritic size is established prior to adolescence and then remains relatively constant into adulthood due to a homeostatic balance between growth and retraction pathways. However, schizophrenia is characterized by accelerated reductions of cerebral cortex gray matter volume and onset of clinical symptoms during adolescence, with reductions in layer 3 pyramidal neuron dendritic length, complexity, and spine density identified in multiple cortical regions postmortem. Nogo receptor 1 (NGR1) activation of the GTPase RhoA is a major pathway restricting dendritic growth in the cerebral cortex. We show that the NGR1 pathway is stimulated by OMGp and requires the Rho guanine nucleotide exchange factor Kalirin-9 (KAL9). Using a genetically encoded RhoA sensor, we demonstrate that a naturally occurring missense mutation in Kalrn, KAL-PT, that was identified in a schizophrenia cohort, confers enhanced RhoA activitation in neuronal dendrites compared to wild-type KAL. In mice containing this missense mutation at the endogenous locus, there is an adolescent-onset reduction in dendritic length and complexity of layer 3 pyramidal neurons in the primary auditory cortex. Spine density per unit length of dendrite is unaffected. Early adult mice with these structural deficits exhibited impaired detection of short gap durations. These findings provide a neuropsychiatric model of disease capturing how a mild genetic vulnerability may interact with normal developmental processes such that pathology only emerges around adolescence. This interplay between genetic susceptibility and normal adolescent development, both of which possess inherent individual variability, may contribute to heterogeneity seen in phenotypes in human neuropsychiatric disease.

Opposing Effects of Cyclooxygenase-2 (COX-2) on Estrogen Receptor ß (ERß) Response to 5α-Reductase Inhibition in Prostate Epithelial Cells.

Current pharmacotherapies for symptomatic benign prostatic hyperplasia (BPH), an androgen receptor-driven, inflammatory disorder affecting elderly men, include 5α-reductase (5AR) inhibitors (i.e. dutasteride and finasteride) to block the conversion of testosterone to the more potent androgen receptor ligand dihydrotestosterone. Because dihydrotestosterone is the precursor for estrogen receptor ß (ERß) ligands, 5AR inhibitors could potentially limit ERß activation, which maintains prostate tissue homeostasis. We have uncovered signaling pathways in BPH-derived prostate epithelial cells (BPH-1) that are impacted by 5AR inhibition. The induction of apoptosis and repression of the cell adhesion protein E-cadherin by the 5AR inhibitor dutasteride requires both ERß and TGFß. Dutasteride also induces cyclooxygenase type 2 (COX-2), which functions in a negative feedback loop in TGFß and ERß signaling pathways as evidenced by the potentiation of apoptosis induced by dutasteride or finasteride upon pharmacological inhibition or shRNA-mediated ablation of COX-2. Concurrently, COX-2 positively impacts ERß action through its effect on the expression of a number of steroidogenic enzymes in the ERß ligand metabolic pathway. Therefore, effective combination pharmacotherapies, which have included non-steroidal anti-inflammatory drugs, must take into account biochemical pathways affected by 5AR inhibition and opposing effects of COX-2 on the tissue-protective action of ERß.

Age-dependent increase in Kalirin-9 and Kalirin-12 transcripts in human orbitofrontal cortex.

KALRN (KAL) is a Rho GEF that is highly involved in regulation of the actin cytoskeleton within dendrites. There are several isoforms of the protein that arise from differential splicing of KALRN's 66 exons. KAL isoforms have different functions in development. For example, overexpression of the KAL9 and KAL12 isoforms induce dendritic elongation in early development. However, in mature neurons KAL9 overexpression reduces dendritic length, a phenotype also observed in normal human ageing. We therefore hypothesized that KAL9 would have increased expression with age, and undertook to evaluate the expression of individual KALRN exons throughout the adult lifespan. Postmortem human brain grey matter from Brodmann's area (BA) 11 and BA47 derived from a cohort of 209 individuals without psychiatric or neurodegenerative disease, ranging in age from 16 to 91 years, were analysed for KALRN expression by Affymetrix exon array. Analysis of the exon array data in an isoform-specific manner, as well as confirmatory isoform-specific qPCR studies, indicated that the longer KAL9 and KAL12 isoforms demonstrated a statistically significant, but modest, increase with age. The small magnitude of the age effect suggests that inter-individual factors other than age likely contribute to a higher degree to KAL9 and KAL12 expression. In contrast to KAL9 and KAL12, global KALRN expression did not increase with age. Our work suggests that global measures of KALRN gene expression may be misleading and future studies should focus on isoform-specific quantification.

Neural deficits in a mouse model of PACS1 syndrome are corrected with PACS1- or HDAC6-targeting therapy.

PACS1 syndrome is a neurodevelopmental disorder (NDD) caused by a recurrent de novo missense mutation in PACS1 (p.Arg203Trp (PACS1R203W)). The mechanism by which PACS1R203W causes PACS1 syndrome is unknown, and no curative treatment is available. Here, we use patient cells and PACS1 syndrome mice to show that PACS1 (or PACS-1) is an HDAC6 effector and that the R203W substitution increases the PACS1/HDAC6 interaction, aberrantly potentiating deacetylase activity. Consequently, PACS1R203W reduces acetylation of α-tubulin and cortactin, causing the Golgi ribbon in hippocampal neurons and patient-derived neural progenitor cells (NPCs) to fragment and overpopulate dendrites, increasing their arborization. The dendrites, however, are beset with varicosities, diminished spine density, and fewer functional synapses, characteristic of NDDs. Treatment of PACS1 syndrome mice or patient NPCs with PACS1- or HDAC6-targeting antisense oligonucleotides, or HDAC6 inhibitors, restores neuronal structure and synaptic transmission in prefrontal cortex, suggesting that targeting PACS1R203W/HDAC6 may be an effective therapy for PACS1 syndrome.

RNA-targeted therapy corrects neuronal deficits in PACS1 syndrome mice.

Neurodevelopmental disorders (NDDs) are frequently associated with dendritic abnormalities in pyramidal neurons that affect arbor complexity, spine density, and synaptic communication 1,2. The underlying genetic causes are often complex, obscuring the molecular pathways that drive these disorders 3. Next-generation sequencing has identified recurrent de novo missense mutations in a handful of genes associated with NDDs, offering a unique opportunity to decipher the molecular pathways 4. One such gene is PACS1, which encodes the multi-functional trafficking protein PACS1 (or PACS-1); a single recurrent de novo missense mutation, c607C>T (PACS1R203W), causes developmental delay and intellectual disability (ID) 5,6. The processes by which PACS1R203W causes PACS1 syndrome are unknown, and there is no curative treatment. We show that PACS1R203W increases the interaction between PACS1 and the α-tubulin deacetylase HDAC6, elevating enzyme activity and appropriating control of its posttranscriptional regulation. Consequently, PACS1R203W reduces acetylation of α-tubulin and cortactin, causing the Golgi to fragment and enter developing neurites, leading to increased dendrite arborization. The dendrites, however, are beset with diminished spine density and fewer functional synapses, characteristic of ID pathology. Treatment of PACS1 syndrome mice with PACS1- or HDAC6-targeting antisense oligonucleotides restores neuronal structure and synaptic transmission, suggesting PACS1R203W/HDAC6 may be targeted for treating PACS1 syndrome neuropathology.

More than a marker: potential pathogenic functions of MAP2.

Microtubule-associated protein 2 (MAP2) is the predominant cytoskeletal regulator within neuronal dendrites, abundant and specific enough to serve as a robust somatodendritic marker. It influences microtubule dynamics and microtubule/actin interactions to control neurite outgrowth and synaptic functions, similarly to the closely related MAP Tau. Though pathology of Tau has been well appreciated in the context of neurodegenerative disorders, the consequences of pathologically dysregulated MAP2 have been little explored, despite alterations in its immunoreactivity, expression, splicing and/or stability being observed in a variety of neurodegenerative and neuropsychiatric disorders including Huntington's disease, prion disease, schizophrenia, autism, major depression and bipolar disorder. Here we review the understood structure and functions of MAP2, including in neurite outgrowth, synaptic plasticity, and regulation of protein folding/transport. We also describe known and potential mechanisms by which MAP2 can be regulated via post-translational modification. Then, we assess existing evidence of its dysregulation in various brain disorders, including from immunohistochemical and (phospho) proteomic data. We propose pathways by which MAP2 pathology could contribute to endophenotypes which characterize these disorders, giving rise to the concept of a "MAP2opathy"-a series of disorders characterized by alterations in MAP2 function.

Investigating Post-translational Modifications in Neuropsychiatric Disease: The Next Frontier in Human Post-mortem Brain Research.

Gene expression and translation have been extensively studied in human post-mortem brain tissue from subjects with psychiatric disease. Post-translational modifications (PTMs) have received less attention despite their implication by unbiased genetic studies and importance in regulating neuronal and circuit function. Here we review the rationale for studying PTMs in psychiatric disease, recent findings in human post-mortem tissue, the required controls for these types of studies, and highlight the emerging mass spectrometry approaches transforming this research direction.

A Schizophrenia-Linked KALRN Coding Variant Alters Neuron Morphology, Protein Function, and Transcript Stability.

BACKGROUND: Large-scale genetic studies have revealed that rare sequence variants, including single nucleotide variants (SNVs), in glutamatergic synaptic genes are enriched in schizophrenia patients. However, the majority are too rare to show any association with disease and have not been examined functionally. One such SNV, KALRN-P2255T, displays a penetrance that greatly exceeds that of previously identified schizophrenia-associated SNVs. Therefore, we sought to characterize its effects on the function of kalirin (Kal)-9, a dual Ras-related C3 botulinum toxin substrate 1 and Ras homologue gene family, member A (RhoA) guanine nucleotide exchange factor, upregulated in human schizophrenia brain tissue. METHODS: Kal9 was overexpressed in primary rat cortical neurons or human embryonic kidney 293 (HEK293) cells. The effects of the P2255T variant on dendritic branching, dendritic spine morphology, protein and messenger RNA stability, and catalytic activity were examined. RESULTS: Kal9-P2255T leads to diminished basal dendritic branching and dendritic spine size, compared with wild-type Kal9. The P2255T SNV directly affected Kal9 protein function, causing increased RhoA activation in HEK293 cells, but had no effect on Ras-related C3 botulinum toxin substrate 1 activation. Consistent with human postmortem findings, we found that Kal9-P2255T protein levels were higher than those of wild-type Kal9 in neurons. Increased messenger RNA stability was detected in HEK293 cells, indicating that this was the cause of the higher protein levels. When analyzed together, increased intrinsic RhoA guanine nucleotide exchange factor catalytic activity combined with increased messenger RNA expression led to net enhancement of RhoA activation, known to negatively impact neuronal morphology. CONCLUSIONS: Taken together, our data reveal a novel mechanism for disease-associated SNVs and provide a platform for modeling morphological changes in mental disorders.

Isolated sinus tachycardia following reinitiation of risperidone in a patient with suspected autonomic hypersensitivity.

The second generation antipsychotic risperidone is generally considered to have low cardiac adverse events, with an increased risk of ventricular arrhythmias being reported only rarely in literature. We report here the case of a patient with a significant history of alcohol dependence, yet with no previous cardiac history, who had previously tolerated risperidone well, but had experienced isolated sinus tachycardia in the post detox period, following the reinitiation of risperidone therapy. The Naranjo Adverse Drug Reaction (ADR) probability scale rating for this being a medication adverse event (AE) was 4, thus indicating that this patient's AE was associated with risperidone therapy. This case report will contribute to the limited evidence of adverse cardiac events associated with risperidone therapy, with particular emphasis on the susceptibility of patients in a state of autonomic hypersensitivity.